Using engineered transcription factors to regulate cancer-related genes has many useful applications, including (i) testing target genes for oncogenicity either through their repression or activation; (ii) validating proposed gene interaction pathways leading to cellular malignancy; and (iii) charactering cancer cell lines by examining the effects of target gene control on either the maintenance or reversal of the cancerous phenotype. In this proposal, we will use rationally-designed zinc finger proteins (ZFPs) to target repressor proteins to the promoter regions of a tumor suppressor protein (p19ARF) and a human oncogene (cyclin D1) (phase I) and demonstrate the effects of chromosomal gene repression on cellular malignancy (Phase II). These experiments will serve to validate our approach as a general method for analyzing cancer genes and lead to the creation of a 'toolbox' of gene repressors for use in cancer research (Phase II). PROPOSED COMMERCIAL APPLICATIONS: These designed transcription factors fulfill an immediate need for cancer gene target validation in drug discovery and have long-term potential in gene therapy.